Attachment sites of carbohydrate moieties to peptide chain of bovine k-casein from normal milk.

Abstract

The attachment site of the carbohydrate moiety to the peptide chain of normal K-casein was investigated with K-casein component P-6 containing the most carbohydrates. Three short glycopeptides 6-IB1, 6-IB2-1, and 6-IB2-2 were prepared by cyanogen bromide cleavage and digestion of proteases (pronase P and thermolysin). Glycopeptides 6-IB1, 6-IB2-1, and 6-IB2-2 corresponded to residues 128_??_139 (Gly-Glu-Pro-Thr-Ser-Thr-Pro-Thr-Thr-Glu-Ala-Val), residues 128_??_132 (or 127_??_131) and residues 135_??_139 of K-casein A, respectively, and contained threonine and or serine, but not asparagine. Glycopeptide 6-IB1 was considered to have three carbohydrate chains because it contained three galactosamine residues. The results of alkali treatment of 6-IB1, under reduction condition excluded serine residue as the binding site, and confirmed the existence of three binding sites in the carbohydrate moieties. The carbohydrate moiety was shown to attach to threonine residue No. 131 from analysis of 6-IB2-1 and to threonine residue No. 135 (or 136) from analysis of 6-IB2-2. It was concluded that the carbohydrate chains attached to threonine residues No. 131, 133 and 135 (or 136).